Infectious agents, such as viruses, bacteria, fungi, or protozoa, encode or carry their own crucial enzymes and nucleic acids which are obvious targets for intervention.
The viruses can be generally classified as being single stranded or double stranded. All viruses with a DNA genome except Pavoviridae are double stranded. Those viruses having RNA genomes are single or double stranded.
As a matter of convention, the nucleotide sequences found in viral messenger RNAs during replication are designated as having positive polarities. This allows RNA viruses to be classified as: 1) negative-stranded (-polarity) viruses, if they have genomes with nucleotide sequences complementary to those present in mRNAs; 2) positive-stranded (+polarity) viruses, if they have genomes with identical sequences to mRNA; and 3) double-stranded RNA viruses, if they have RNA strands of both polarities. In addition to the polarity differences between viral RNA genomes, these viruses can be distinguished based on the segmentation of viral RNA.
The polarity of the RNA genome helps determine the mechanism of viral replication. Genomes of RNA viruses having single stranded RNAs of plus polarity are directly translated during the initial stages of infection by cell macromolecules without requiring the synthesis of specific viral transcriptases and other viral enzymes necessary for mRNA formation. Negative- and double-stranded viruses, however, require that transcription of mRNA from genome RNA occurs before translation can be initiated. Since normal cells lack an enzyme that transcribes RNA templates into complementary RNA strands, these viruses must encode the appropriate enzyme(s) during their replication cycle. Furthermore, the enzyme must be incorporated into progeny virus in order to initiate transcription and replication during successive infections.
The life cycle of retroviruses requires a specific protease that processes the precursor "gag" and "pol" polyproteins into mature virion components. If the protease is absent or inactive, non-infectious virus particles are produced.
All replicative competent viruses contain a minimum of three nucleotide sequence stretches that can be transcribed into protein without encountering a stop codon. These open reading frames are 1) "gag", which encodes virion core structural proteins; 2) "pol", which encodes the enzymes that catalyze the synthesis of viral DNA and its integration into host DNA; and 3) "env", which encodes the glycoproteins contained in the external and transmembrane virion envelope.
The retroviral life cycle begins when viral envelope proteins attach to specific receptors on the surface of susceptible cells. Reverse transcription of the single-stranded viral RNA genome(s) in the cytoplasm of infected cells leads to the formation of DNA-RNA heteroduplex consisting of the RNA plus strand and a DNA minus strand. The single stranded DNA then serves as a template, allowing the polymerase activity of reverse transcriptase to make a second DNA copy (the plus strand), whose sequence corresponds to that of the RNA contained in the core of the virus. The minus DNA strand is synthesized in a continuous fashion from a RNA primer, whereas the plus strand is synthesized discontinuously using multiple initiation sites. The viral genetic information now in the form of a double stranded, linear molecule or single strand linear molecule migrates to a cell nucleus where it is integrated into the host-cell genome. Following integration the viral DNA will be copied along with the cell's own genes every time the cell divides.
As the new virus comes from the host cells, one of the enzymes contained within the precursor polyprotein(s) cleaves itself from the precursor. This precursor is cleaved to produce the mature proteins, by the viral protease, which is itself a part of the precursor and becomes activated when the new progeny particles leave the surface of the infected cell.
Viruses which comprise negative-single stranded RNA genomes include Orthxomyoviridae, Rhabdoviridae, Paramyxoviridae, Bunyaviridae, and Arenaviridae.
Orthomyxoviridae, Bunyaviridae and Arenaviridae have segmented while Rhabdoviridae and Paramyxoviridae have unsegmented genomes.
Bunyaviridae include members of the Phlebovirus and Rotavirus which infects mammals, and the plant reoviruses, Phytoreovirus and Fijivirus.
Orthomyxoviridae includes the influenza viruses A, B and C.
Rhabdoviridae viruses are found in vertebrates as well as in plants. The genera Lyssavirus includes rabies virus and Vesiculovirus (vesicula stomatitis virus and chandipura virus).
Paramyxoviridae includes parainfluenza virus of mammals (mumps virus, Newcastle disease virus, etc.), Morbillivirus (the viruses of measles, canine distemper, etc.) and Pneumovirus (respiratory syctytial viruses of man and cattle).
Viruses which comprise positive single stranded RNA genomes include Picornaviridae, and Togaviridae.
Picornaviridae genera includes Enterovirus (Polioviruses, (Coxsackievirsus, Enterovirus 72 (hepatitis A), Cardiovirus (Encephalomyocarditis) Rhinovirus and Aphthovirus (foot and mouth disease of cattle).
Togaviridae genus includes Alphavirus (Sindbis virus, Semliki Forest virus), Flavivirus (Yellow fever virus, dengue virus, tick borne virus), Rubivirus (Rubella virus) and Pestivirus (Micosal disease virus).
Parvovirus is the only virus having a single stranded negative DNA genome. This virus primarily infects cats and dogs.
All the virus with a DNA genome with the exception of Pavoviridae are double stranded. These viruses include Papovaviradae, Adenoviridae, Herpesviridae, Iridoviridae, Poxviridae and Hepadnovidae.
Papovaviradae includes the genera Papillomavirus which infects humans with papilloma or wart virus and SV-40 like virus.
Adenoviridae comprises the two genus Mastadenovirus and Aviadenovirus of which Mastadenovirus infects mammals.
Herpesviridae and Alphaherpesvirinae include simplexvirus (herpes simplex types 1 and 2 which infects humans and Bovine mammillitis virus), Poikilovirus (pseudorabies virus) and Varicellavirus (human herpesvirus 3).
Hepadnaviridae includes the genus hepadnavirus (human hepatitis B virus, and mammalian hepatitis virus).
Poxviridae includes the genus orthopoxvirus which infects humans and animal poxvirus such as parapoxvirus and capripoxvirus.
U.S. Pat. No. 4,496,689, which is herein incorporated by reference, discloses conjugates of heparin, PVA or PEG (polyethylene glycol) which have active sites and can be used in the present invention.
U.S. Pat. No. 5,134,119 to Lezdey et al which is herein incorporated by reference, discloses the analogs of alpha 1-antitrypsin which can be used in the present invention.
U.S. Pat. No. 5,217,951 to Lezdey et al, which is herein incorporated by reference discloses the analogs, salts and derivatives of serine protease inhibitors for use in the treatment of non-bronchial mast cell implicated diseases.
U.S. Pat. No. 4,496,689 to Mitra, which is herein incorporated by reference discloses the preparation of complexes or conjugates of alpha 1-antitrypsin which can be used in the present invention.
Mammalian proteinase inhibitors are classified into families called kunins, kazals, ALPs (antileukoproteases), serpins, .alpha.-macroglobulins, cystatins and TIMPs (Tissue Inhibitors of Metalloproteinase).
Kunins include aprotinin, trypstatin and inter -.alpha.- trypsin inhibitor.
Serpins include alpha 1-antitrypsin inhibitor, alpha 1-antichymotrypsin, antithrombin, C 1-inhibitor, alpha 2- antiplasmin and Protein C-inhibitor.
Alpha 2-macroglobulin, although not a serpin, acts similarly to both alpa 1-antitrypsin and alpha 1-antichymotrypsin. This compound is considered as a scavenger which binds with those nucleotide groups, including aspartic groups, that are not picked up by the serpins.
Alpha 1-antichymotrypsin is considered by many as being misnamed since this inhibitor does not primarily bind with chymotrypsin but rather cathepsin G. Alpha 1-antitrypsin is the inhibitor with a preference for chymotrypsin as a binding partner.
It is to be understood that the term "conjugate" as used herein also refers to the complexes which may be formed, for example, with a polysaccharide polyol or protease (cathepsin G, elastase, etc) wherein the active sites of the protease inhibitor are retained.